1. Field of the Invention The present invention relates to scanning optical microscopes and, more particularly, to a laser scanning microscope (LSM) that performs focal point movement along the direction of the optical axis by using a wavefront converting element.
2. Discussion of Related Art
It has heretofore been necessary in order to obtain a three-dimensional image of a specimen with an LSM, for example, to capture optical images of successive planes inside the specimen by mechanically moving either the specimen or the objective along the direction of the optical axis. With this method, however, it is difficult to realize positional control with high accuracy and high reproducibility because the method needs a mechanical drive. In a case where the specimen is moved, high-speed scanning cannot be effected when the size of the specimen is large.
In observation of a biological specimen, if the objective is-scanned in the state of being in direct contact with the specimen or immersed in a culture solution of the specimen, vibrations of the objective adversely affect the specimen under observation.
To solve the above-described problems, Japanese Patent Application Unexamined Publication (KOKAI) No. Hei 11-101942 discloses an adaptive optical apparatus. The apparatus is a microscope having an optical element (wavefront converting element) capable of changing power. The arrangement of the microscope is shown in FIGS. 27 and 28. In this prior art, a wavefront converting element is inserted in either or both of a viewing optical path and an illuminating optical path to change the focal length of the optical system and to correct aberration due to the change of the focal length by using the wavefront converting element. With this arrangement, it is possible not only to form and move a focal point in the object space without changing the distance between the objective and the specimen but also to correct aberration.
In the above-described prior art, it is preferable to place the wavefront converting element in the pupil plane of the objective or at a position conjugate to the pupil plane from the viewpoint of allowing the wavefront converting element to effectively perform its functions of moving the focal point in the object space and making aberration correction. If the wavefront converting element is not conjugate to the pupil plane, illuminating light or image-forming light will pass at different positions on the wavefront converting element according to the height of the object detected by the objective. To perform focal point movement or aberration correction, the wavefront shape has to be changed according to the object height. If the wavefront shape cannot properly be changed, image quality is likely to degrade considerably in an area where the object height is high.
If the wavefront converting element is changed into an optimum shape in accordance with a change in the object height, even if the wavefront converting element is not conjugate to the pupil plane, it is possible to avoid image quality degradation in an area where the object height is high. To realize this, however, the wavefront converting element needs to be controlled at high speed so as to provide an optimum rotationally asymmetric configuration. This is extremely difficult.
For the reasons stated above, it is desirable that the wavefront converting element should be placed at a position conjugate to the pupil. This is, however, difficult to implement because of the following problems.
A variety of objectives are used in microscopic observation, and the pupil position differs for each objective. Therefore, when a plurality of objectives are switched from one to another to perform observation, it is difficult to keep the pupils of the objectives in conjugate relation to the wavefront converting element at all times.
Further, the wavefront convening element needs to be placed in conjugate relation to the position of a laser scanning member and also to the position of the objective pupil. Accordingly, at least two pupil relay optical systems are required. Therefore, the size of the apparatus becomes large and it becomes unfavorably complicated.
Further, in the above-described prior art, a reflection type wavefront convening element is incorporated in the illuminating optical path and/or the light-detecting optical path. Therefore, the prior art uses beam splitters as shown in FIGS. 27 and 28. Accordingly, when a non-polarized laser is used as a light source, together with a non-polarization type beam splitter, the amount of light is reduced to xc2xc every time the laser beam travels via the wavefront converting element.
More specifically, the amount of light is reduced to xc2xcin the process of illumination and also reduced to xc2xcin the process of detection. That is, the amount of light is reduced to {fraction (1/16)}in total. If a linearly polarized laser is used as a light source, together with a polarization beam splitter and a quarter-wave plate, the loss of light in the process of illumination can be prevented. However, in observation of fluorescence in a non-polarized state, the amount of light is reduced to xc2xdin the process of (fluorescence) detection.
Further, even when a polarization beam splitter and a quarter-wave plate are used as stated above, it is not always possible to use a linearly polarized laser as a light source. If a non-polarized laser is used to observe fluorescence, the amount of light is reduced to xc2xdin the process of illumination and also reduced to xc2xdin the process of detection. That is, the amount of light is reduced to xc2xcin total.
The present invention was made to solve the above-described problems associated with the prior art. Objects of the present invention are as follows. A first object of the present invention is to provide a scanning optical microscope, e.g. a laser scanning microscope (LSM), using a wavefront converting element, wherein even when the object pupil and the wavefront converting element are not placed in conjugate relation to each other, off-axis performance degradation is minimized, and wherein the wavefront converting element can be controlled by an extremely simple method, and a pupil relay optical system is simple in arrangement or unnecessary. A second object of the present invention is to provide an LSM using a wavefront converting element, in which the loss of light can be prevented even when the wavefront converting element is of the reflection type.
To attain the above-described objects, the present invention provides a first scanning optical microscope including a light source and a wavefront converting element for applying a desired wavefront conversion to illuminating light emitted from the light source. An objective collects wavefront-converted illuminating light emerging from the wavefront converting element onto a sample. A detector detects signal light emitted from the sample. An actuator scans the objective along a direction perpendicular to the optical axis.
It is desirable that illuminating light emerging from the wavefront converting element should be an approximately parallel beam.
In addition, the present invention provides a second scanning optical microscope wherein when the above-described actuator scans one section of the sample perpendicular to the optical axis with the objective, the wavefront converting element applies a constant wavefront conversion to the illuminating light.
In addition, the present invention provides a third scanning optical microscope having an arrangement similar to that of the first or second scanning optical microscope, wherein when the amount of movement of the objective along the direction perpendicular to the optical axis (this will hereinafter be referred to as xe2x80x9cscan rangexe2x80x9d) is denoted by xcex94X, the following condition (1) is satisfied:
xcex94Xxe2x89xa60.66fORxc2x7xcex(xcex94Xxc2x7NA4) xe2x80x83xe2x80x83(1) 
where:
fOB: the focal length of the objective;
xcex94Z: the amount of focal point movement caused by the wavefront converting element;
xcex: the wavelength of the illuminating light;
NA: the numerical aperture of the objective.
In addition, the present invention provides a fourth scanning optical microscope including a light source and an optical element having a positive power for converting illuminating light emitted from the light source into a convergent beam. The fourth scanning optical microscope further includes a reflecting mirror with an aperture and a reflection type wavefront converting element for applying a desired wavefront conversion to the illuminating light. An objective collects the wavefront-converted illuminating light onto a sample. A detector detects signal light emitted from the sample.
In addition, the present invention provides a fifth scanning optical microscope wherein an optical system including the reflecting mirror with an aperture in the fourth scanning optical microscope satisfies the following condition (2):
rHmin/rMincxe2x89xa60.5 xe2x80x83xe2x80x83(2) 
where:
rHmin: the minimum value of the length from the optical axis to the reflecting mirror edge;
rMinc: the radius of wavefront-converted illuminating light incident on the reflecting mirror with an aperture.
In addition, the present invention provides a sixth scanning optical microscope including a light source and an optical element having a positive power for converting illuminating light emitted from the light source into a convergent beam. A reflecting mirror is placed at a position where the convergent beam is collected. A reflection type wavefront converting element applies a desired wavefront conversion to the illuminating light. An objective collects the wavefront-converted illuminating light onto a sample. A detector detects signal light emitted from the sample.
In addition, the present invention provides a seventh scanning optical microscope wherein an optical system including the reflecting mirror in the sixth scanning optical microscope satisfies the following condition (3):
rMmin/rAincxe2x89xa60.5 xe2x80x83xe2x80x83(3) 
where:
rMmin: the minimum value of the length from the optical axis to the reflecting mirror edge;
rAinc: the radius of wavefront-converted illuminating light at the position of the reflecting mirror.
In addition, the present invention provides an eighth scanning optical microscope including a light source and a reflection type wavefront converting element for applying a desired wavefront conversion to illuminating light emitted from the light source. An objective collects wavefront-converted illuminating light onto a sample. The light source also serves as a detector for detecting signal light emitted from the sample.
In addition, the present invention provides a ninth scanning optical microscope including a light source and a reflection type wavefront converting element for applying a desired wavefront conversion to illuminating light emitted from the light source. An objective collects wavefront-converted illuminating light emerging from the wavefront converting element onto a sample. A detector detects signal light emitted from the sample. The reflection type wavefront converting element is placed in an optical path so as to satisfy the following condition (4):
xcex8PRxe2x89xa650xc2x7NAxe2x88x921(xcexxc2x7xcex94Zxe2x88x921)xe2x80x83xe2x80x83(4) 
where:
xcex8PR: the angle (xc2x0) of incidence of the principal ray on the wavefront converting element;
xcex94Z: the amount of focal point movement;
xcex: the wavelength of the illuminating light;
NA: the numerical aperture of the objective.
In addition, the present invention provides a tenth scanning optical microscope including a light source and a reflection type wavefront converting element for applying a wavefront conversion to illuminating light emitted from the light source. An objective collects wavefront-converted illuminating light emerging from the wavefront converting element onto a sample. A detector detects signal light emitted from the sample. The reflecting surface of the reflection type wavefront converting element is controllable into an aspherical toric surface configuration.
Still other objects and advantages of the invention will in part be obvious and will in part be apparent from the specification.
The invention accordingly comprises the features of construction, combinations of elements, and arrangement of parts which will be exemplified in the construction hereinafter set forth, and the scope of the invention will be indicated in the claims.